The Spectrin and actin network on the cytoplasmic side of the red cell membrane is believed to be an important determinant of cell deformability and osmotic fragility. Recent work done in our laboratory showed that a supramolecular complex made of spectrin, actin and band 4.1 acted as nuclei for the polymerization of actin. Under appropriate conditions, the complex causes gelation of F-actin or form large aggregate with spectrin which can be sedimented by low-speed centrifugation. We plan to use both biochemical and biophysical approaches to study the structure and function of the spectrin and actin network in relation to cell age and hope to correlate the well known phenomenon of decreased deformability and increased susceptibility to osmotic lysis in older cells to abnormality in the structure and function of the contractile-protein network. The biochemical approach will be on the characterization of the contractile proteins in the intact cell and in isolated form, including studies on the degree of phosphorylation, state of aggregation or crosslinking, sensitivity to drugs that are known to affect contractile proteins (e.g. cytochalasins). The biophysical approach involves the measurement of the fluidity of the lipid region of the membrane, using electron paramagnetic resonance techniques, in cells of different age and under various experimental conditions. In addition, the physical properties of spectrin and actin from cells of different age will be studied with viscometric and light scattering techniques. With these studies, we hope to establish a correlation between the state of the contractile proteins and the age-related rigidity of the red cell membrane.